Nuclear factor-κ B (NFκB) signaling is an essential signal transduction pathway involved in inflammatory responses, oncogenesis, viral infection, the regulation of cell proliferation and apoptosis and, in particularly in the case of B and T lymphocytes, in antigenic stimulation (Ghosh, 1998, Annu. Rev. Immunol., 16, 225-260; Karin, 1999, J. Biol. Chem., 274, 27339-27342; Israel, 2000, Trends Cell. Biol., 10, 129-133; Santoro, 2003, EMBO J., 22, 2552-2560). In mammalian cells, there are five NFκB family members that dimerize: RelA, RelB, c-Rel, NFκB 2/p100/p52 and NFκB 1/p105/p50. NFκB, whose predominant form is a heterodimeric transcription factor composed of p50 and RelA subunits, remains sequestered in the cytoplasm through association with members of an inhibitory family of proteins known as IκB. Upon stimulation by the cytokines TNF-α and interleukin-1, endotoxin (LPS), microbial and viral infections, pro-inflammatory signals converge on the canonical IkB kinase complex (IKK), a protein complex that is composed of two kinase subunits, IKKα/IKK-1 and IKKβ/IKK-2 and a structural/regulatory subunit NEMO/IKK-γ. Once activated IKK complex phosphorylates IkB proteins, triggering their ubiquitination and subsequent degradation by the proteasome. Free NFκB can then move into nucleus to initiate or up-regulate gene expression.
Although IKKα and IKKβ exhibit striking structural similarity (52%), genetic studies have shown that they are involved in two pathways for the activation of NFκB (Pomerantz, 2002, Molecular Cell 2002 10: 693-695). IKKβ has been identified as the pro-inflammatory kinase responsible of activation of classical NFκB complexes, whereas IKKα in association with NFκB inducing kinase (NIK) plays an essential role in the non-canonical NFκB signaling pathway (Senftleben, 2001, 293: 1495-1499).
NFκB plays an essential role in the development and progression of cancer, including breast cancer. Animal studies suggest the presence of constitutively active NFκB at an early stage during neoplastic transformation of mammary cells (Clarkson et al., 2000, J Bio Chem. 275(17):12737-42). NFκB inhibits apoptosis in mouse mammary epithelia (Sovak et al., 1999, Cell Growth Differ. 10(8):537-44) and selective activation of NFκB subunits have been found in human breast cancer cell lines and patient samples (Sovak et al., 1997, J Clin Invest. 100(12):2952-60; Cogswell et al., 2000, Oncogene 19(9):1123-31). An inverse correlation between the levels of NFκB activation and estrogen receptor expression has been reported (Nakshatri et al., 1997, Mol Cell Biol. 17(7):3629-39) and inhibition of NFκB in breast cancer cells induces spontaneous apoptosis (Sovak et al., 1999, Cell Growth Differ. 10(8):537-44; Cogswell et al., 2000, Oncogene 19(9):1123-31). Paclitaxel-induced sensitivity of breast cancer cell lines was enhanced by an NFκB inhibitor, parthenolide (Patel et al., 2000, Oncogene 19(36):4159-69; Newton et al., 1999, J Bio Chem. 274(26):18827-35). The Mullerian inhibiting substance was also found to inhibit breast cancer growth through NFκB mediated pathway (Segev et al., 2000, J Bio Chem. 275(37):28371-9). Furthermore, the transactivation function of NFκB is negatively regulated by IκBβ1 in breast cancer cell lines (Newton et al., 1999, J Bio Chem. 274(26):18827-35). Lastly, overexpression of HER2/neu can activate NFκB through the activation of Akt pathway and block apoptosis (Zhou et al., 2000, J Bio Chem. 275(10:8027-31). All these reports together suggest that NFκB plays an important role in cancer generally and in breast cancer specifically.
In light of the foregoing, inhibition of NFκB activation represents a target for development of new anti-inflammatory and anti-cancer drugs (Poulaki, 2002, Am J Pathol. 161: 2229-2240). Among many protein actors in NFκB signaling pathway, IKK complex represents one of the most promising molecular targets for discoveries of new specific NFκB inhibitors. To minimize the potential toxicity effects in vivo, therapeutic success will greatly depend on the abilities of the NFκB inhibitors to block activating signals without modifying the basal level of NFκB activity. For example, May et al. described a cell-permeable peptidic inhibitor that specifically blocks the pro-inflammatory NFκB activation by disrupting the constitutive NEMO interaction with IKK kinases (May, 2000, Science 289, 1550-1554; May, 2002, J. Biol. Chem. 277, 45992-46000). Accordingly, it would be desirable to identify additional small molecules that can reduce NFκB activity by selectively inhibiting NFκB and/or components of the NFκB signaling pathway.